The very low levels of endogenous DNA remaining in most ancient specimens has precluded the shotgun sequencing of many interesting samples due to cost. For example, ancient DNA (aDNA) libraries derived from bones and teeth often contain <1% endogenous DNA, meaning that the majority of sequencing capacity is taken up by environmental DNA. Thus much of the cost associated with sequencing low endogenous DNA sample provides no human genome data. As a result, many ancient DNA samples are considered unsuitable for sequencing because the data yield is low compared to the resources required. Thus there is a need in the art to increase endogenous DNA yield in low endogenous DNA samples and specifically to increase the percent of endogenous DNA being sequenced when sequencing low endogenous DNA samples.
Recent developments in DNA extraction have provided lower cost next-generation sequencing techniques to the point that the field of paleogenetics has transitioned from focusing on PCR-amplified mitochondrial DNA and Y-chromosomal markers to shotgun sequencing of the whole genome. However, shotgun sequencing can yield less than desirable results when sequencing low endogenous DNA samples due to the low percentage of endogenous DNA in the overall sample material.
Instead, the use of autosomal DNA sequences may be superior for population genetic analysis because it provides information from both lineages (i.e., maternal and paternal). Thus there exists a specific need in the art to provide an autosomal DNA sequencing technique for ancient DNA analysis in order to derive improved resolution for population genetic analysis. For example, whole genome sequencing of single ancient genomes, including Neanderthals, Denisovan, a Paleo-Eskimo, the Tyrolean Iceman, and an Australian Aborigine, have transformed our understanding of human migrations and revealed previously unknown admixture among ancient populations. However, most of these specimens were exceptional in their levels of preservation: the Neanderthal and Denisovan bones, found in caves, contained ˜1-5% and 70% endogenous DNA, respectively, while the Paleo-Eskimo and Aborigine genomes were obtained from hair specimens, which generally contain lower levels of contamination but are not available in most archaeological contexts.
In contrast, sequencing libraries derived from bones and teeth from temperate environments typically contain <1% endogenous DNA. While samples with 1-2% endogenous DNA can still, with sufficient sequencing, yield enough information for population genetic analyses, the required amount of sequencing of specimens with less DNA is costly and thus untenable for many researchers. Ancient DNA researchers have begun to address this issue by using targeted capture to enrich for only the mtDNA or for a single chromosome. However, due to the highly fragmented nature of ancient DNA, an ideal enrichment technique would extract as much of the endogenous genome as possible so as not to discard any potentially informative sequences. Similar problems exist in forensics.